Fig. 2: Nsp15 prefers unpaired U over paired and mismatched Us, even in substrates without 2′-F modification.
From: Spontaneous base flipping helps drive Nsp15’s preferences in double stranded RNA substrates
A Substrate design. The target strand is labeled with 5′ Cy5 and 3′ Fl, and contains 11 cleavable Us. The complement strand is unlabeled. Three substrates were created with this design by changing the nucleotide complement to U14 to produce either a U-A, U•C, or unpaired U. B Representative denaturing PAGE gels of time-course nuclease assays for each of the three substrates. The U-A and U•C substrates show similar patterns of cleavage with Us in the 4 positions nearest the ends being most favored. The unpaired U substrate shows a different pattern, with cleavage at the unpaired U14 outpacing cleavage at all other Us. Note the near-complete disappearance of the target strand by 30 min, approximately twice as fast as in U-A and U•C substrates. Three independent reactions (each using a distinct protein preparation) were performed with each substrate. Images of each gel are provided in the Source Data file.
